The proposed research addresses a series of questions that emerge from the fact that elderly persons have been exposed to a myriad of pathogens over their lifespan. This immunological history leads, in some cases, to the generation of expanded populations of memory CD8 T cells that have reached the stage of replicative senescence. In cell culture, CD8 T cells that reach this state after repeated rounds of antigen-driven proliferation show irreversible cell cycle arrest, permanent loss of CD28 gene expression, apoptosis resistance, poor response to stress, altered cytokine profiles, and shortened telomeres compared to their CD28+ progenitors. Clinical studies have identified correlations between CD8 T cells showing characteristics of replicative senescence and such diverse health outcomes as reduced responsiveness to influenza vaccination and osteoporotic fractures. The central hypothesis of the proposed research is that the high proportion of senescent CD8 T cells present in vivo exerts deleterious effects on both immune and non-immune organ systems during aging. To further elucidate the underlying mechanisms by which CD8 T cell replicative senescence may mediate pleiotropic physiological effects, the following specific aims will be addressed: (1) To determine the role of senescent CD8 T cells on immune function. Direct cell interaction and cell-free supernatants from senescent cultures will be evaluated for their impact on CD4 T cell help, CD8 effector functions, antigen-presentation and antibody production. (2) To evaluate genetic and non-genetic manipulations to reverse CD8 T cell replicative senescence. Lentivirus vectors containing CD28 or telomerase, reduced levels of oxygen, and exposure to estrogen (which affects several genes associated with senescence) will be compared for effects on telomere/telomerase dynamics, population doublings, CD8 T cell immune functions, and immune modulatory functions identified in Aim 1. (3) To investigate the role of senescent CD8 T cells in osteoporosis, based on the well-documented effects of chronic immune activation on bone integrity and on the identification of cytokine-producing T cells within the bone marrow. Early passage and senescent CD8 T cells will be compared for effects on the maturation, differentiation, and function of osteoclasts (the bone resorbing cells) and osteoblasts (the bone forming cells). In vitro observations will be confirmed using a murine model of osteoporosis. In sum, the proposed studies will provide an unparalleled opportunity to experimentally dissect the genetic and molecular basis of several fundamental aspects of human immunological aging that significantly impact health and longevity.